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243 | P a g e International Standard Serial Number (ISSN): 2319-8141

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International Journal of Universal Pharmacy and Bio Sciences 6(2): March-April 2017

INTERNATIONAL JOURNAL OF UNIVERSAL

PHARMACY AND BIO SCIENCES IMPACT FACTOR 2.96***

ICV 6.16***

Pharmaceutical Sciences RESEARCH ARTICLE …………!!!

VALIDATION OF BUNG PROCESSOR

Shridhar .M*, M.P Gowrav, Gangadharappa .H.V

Department of Pharmaceutics (Pharmaceutical Quality Assurance)

Jagadguru Sri Shivarathhreeswara University, Sri Shivarathreeshwara Nagara, Mysuru-

570015 Karnataka, India.

KEYWORDS:

Validation of autoclave,

Vacuum leak test, Bowie-

dick test, Heat

distribution test, Heat

penetration test.

For Correspondence:

Shridhar .M*

Address:

Department of

Pharmaceutics

(Pharmaceutical Quality

Assurance)

Jagadguru Sri

Shivarathhreeswara

University, Sri

Shivarathreeshwara

Nagara, Mysuru-570015

Karnataka, India.

ABSTRACT

In accordance with GMP, each pharmaceutical company should identify

what qualification work is required to prove that the critical aspects of

their particular operation are controlled. The key elements of a

qualification and validation programme of a company should be clearly

defined and documented. Qualification is the integral part of GMP and

there is no effective QA program without qualification. Now-a-days it is

mandatory to incorporate qualification activity for any system in the

manufacturing premises for all pharmaceutical industries. The purpose of

this study is to initially develop the sterilization process parameter for the

porous load articles then implement the sterilization process for the

porous articles. The process development included qualification of

equipment and the articles. The autoclave cum bung processor which is

used for the cleaning and sterilizing rubber stoppers, garments and

machine parts. Initially installation qualification was done followed by

performing the qualification of the equipment by performing Vacuum

leak test, Bowie dick test, heat distribution test and heat penetration test.

The equipment passes all tests and hence the equipment is suitable for

sterilization purpose as it meets the predetermined specification and

quality attributes. Hence the autoclave cum bung processor is considered

to be qualified.



INTRODUCTION:

An autoclave is a pressure chamber used to carry out industrial processes requiring elevated

temperature and pressure different to ambient air pressure. Autoclaves are used in medical

application to perform sterilization, and in the chemical industry to cure coatings, vulcanize rubber

and for hydrothermal synthesis1.

Autoclave was invented by Charles Chamber land in 18792. The name comes from Greek auto-

self, and Latin clavis - key, a self- locking device3.

The standard temperature and pressure of an autoclave Processes conducted at high temperatures

for short time periods are preferred over lower temperatures for longer times. Some standard

temperatures/pressures employed are 115 °C/10 psi, 121 °C/ 15 psi, and 132 °C/27psi. (psi=pounds

per square inch). In universal autoclave, autoclaving generally involves heating in saturated steam

under a pressure of approximately 15 psi, to achieve a chamber temperature of a least 121 °C (250

°F) but in other applications in industry, for example, other combinations of time and temperature

are sometimes used. Please note that after loading and starting the autoclave, the processing time is

measured after the autoclave reaches normal operating conditions of 121 °C (250 °F) and 15 psi

pressure, not simply from the time you push the "on" button4,5

.

There are four basic types of autoclaves. These types depend upon the method employed to remove

air from the sterilization chamber: Downward displacement autoclaves, Positive pressure

displacement autoclaves, Negative pressure displacement autoclaves and Triple vacuum autoclave.

The downward displacement autoclave is also referred to as a gravity displacement unit. This is

because of the method of air removal in the sterilization chamber. A pool of water in which an

element is immersed, turns to steam when heated. As steam is lighter than air, it forces the air in the

sterilization chamber downward and out through a drain hole. Once the temperature in the

sterilization chamber is sufficient, the drain hole is closed automatically and the sterilization

process begins.

A positive pressure displacement autoclave is an improvement upon the design of a downward

displacement unit. Steam is created in a second, separate chamber and held until the proper amount

to displace all of the air in the sterilization chamber is accumulated. The steam is then released into

the sterilization chamber in a pressurized blast, forcing the air out through the drain hole and

starting the sterilization process.

A negative pressure displacement autoclave is one of the most accurate types available. Once the

sterilization chamber door is closed, a vacuum pump removes the air. Steam is created in a second,

separate chamber. Once the air has been completely removed from the sterilization chamber, the



steam is then released into the sterilization chamber in a pressurized blast much like that of a

positive pressure displacement unit.

A triple vacuum autoclave is set up in a similar fashion to a negative pressure displacement unit in

that there is a vacuum pump to remove air from the sterilization chamber and steam is created in a

second, separate chamber or unit. The process begins by the vacuum removing the air, then a pulse

of steam. This is repeated three times, hence the name "triple vacuum" autoclave6.

The working process of autoclave

Basically, steam enters the chamber jacket, passes through an operating valve and enters the rear of

the chamber behind a baffle plate. It flows forward and down through the chamber and the load,

exiting at the front bottom. A pressure regulator maintains jacket and chamber pressure at a

minimum of 15 psi, the pressure required for steam to reach 121 °C (250 °F).

Overpressure protection is provided by a safety valve. The conditions inside are thermostatically

controlled so that heat (more steam) is applied until 121 °C is achieved, at which time the timer

starts, and the temperature is maintained for the selected time.

Figure 1: The temperature profile for a saturated steam vented cycle of autoclave

tₒ to tn is the time interval in the computation of the Fₒ value. F1 to Fn areas represent the lethal dose

provided to the material being treated. The shaded area under the curve obtained through

summation of F1 to Fn values represents the total Fₒ value. In the heating phase, saturated steam is

admitted into the chamber, displacing the cold air until the exposure temperature and corresponding

saturated steam pressure are attained. In the exposure phase, the sterilizing temperature is

maintained by saturated steam for the prescribed exposure time. The cooling phase can be achieved

by slow exhaust (for containers filled with liquids) or fast exhaust (for goods required to be dry

after sterilization). This phase is completed when the pressure reaches atmospheric pressure7.



F0 = Σ 10 (T-121)/z ∆t Where

F0 = F0 Value

T = Average Temperature

z = Value which indicates relationship of lethality to temperature

= 10 °C (assumed)

121 °C = Sterilization Temperature

“F0” is defined as the number of equivalent minutes of steam sterilization at 121 °C delivered to a

load (product)

D-value (decimal reduction value): it is time required at temperature T to reduce a specific

microbial population by 90% or by a factor of 10

Z-value: it is the number of degrees of temperature change necessary to change the D-value by a

factor of 108.

Industrial and scientific autoclaves

Although best known as sterilizers, autoclaves can also be used to carry out all sorts of industrial

processes and scientific experiments that work best at high-temperatures and pressures. Unlike

sterilizing autoclaves, which usually circulate steam, industrial and scientific autoclaves may

circulate other gases to encourage particular chemical reactions to take place. Industrial autoclaves

are often used for "curing" materials. For example:

Rubber can be vulcanized (heated, toughened, and hardened with sulfur) in an autoclave.

Nylon (a plastic) can be made by "cooking" a concentrated salt solution in an autoclave to

encourage what's called condensation polymerization.

Some autoclaves combine elements of both sterilization and industrial manufacture. For example,

natural cork (wooden) bottle stoppers have to be boiled and sterilized before they're suitable for use.

Traditionally, that was done in large water tanks; now it's much more likely to be done on a large

scale in computer-controlled, industrial autoclaves9.

Validation

The documented evidence that the process, equipment, facilities or systems, operating within

established parameters, can perform effectively and reproducibly giving results meeting

predetermined specifications.

Validation master plan

Describes the areas of activities within which validation is to take place and provides an overview

of the status of planning. It lists the areas, systems and projects being managed, defines the status of

validation for each and gives a broad indication of when validation is to be completed. It is a

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general plan and would normally cover all equipment and processes. It should include all systems

for which validation is planned.

Validation plan

Description of the validation activities, responsibilities and procedures. It describes specifically

how the validation is to be done and details responsibilities if the validation team.

Validation protocol

Outlines the objectives of validation of a specific equipment, testing protocol including elements

such as installation, operational and performance qualification and documentation.

Within the laboratory there should a validation policy which is a strategy document that clearly

defines what the validation process is and its purpose within the laboratory. The policy should make

a commitment to maintaining critical processes and systems in a valid state and should mention

applicable regulations, standards and guidelines that underpin the laboratory’s approach to

validation. The validation policy will specify what should be validated and how validation is

executed as defined in the validation master plan.

The validation master plan (VMP) is a quality management system document. It should be a

controlled document, approved by senior laboratory management and regularly reviewed and, if

necessary, revised in response to organizational and operational changes.

The VMP should be based on and refer to the validation policy. The VMP should clarify

Under what circumstances

Who is responsible

How the validation will be performed and documented

How the validated state will be maintained through regular servicing and calibration and re-

qualification.

For large projects such as the implementation of a new laboratory information system or relocation

of the laboratory to new facilities, it may be appropriate to have a project-specific VMP. The VMP

will inform those working as part of the project team to ensure that the project delivers processes

and systems that are in a validated state and fit for use. Also, it may be desirable for reasons of

complexity to have separate VMPs covering different systems such as computer systems and

automated test systems.

A VMP should cover

The organizational structure and responsibilities for validation activities

Summary or list of equipment, facilities or systems to be validated and qualified

How validation is planned and scheduled



The formats for validation documentation

Links to other relevant quality system processes (e.g .document control, training, equipment

calibration and maintenance.)

Validation process

Each part of the validation process should be documented. There should be a written plan for

performing each validation to specify who is responsible for managing and performing the various

validation tasks such as production of validation protocols and approvals of validation

documentation. Validation protocols should be written for each phase of the validation to include

acceptance criteria. The validation plan and the validation protocols may be combined into a single

document. The outcome of each phase of validation should be recorded and the overall conclusions,

with a scientific assessment of any failures should be documented in a validation summary report.

The validation records and summary report must be reviewed and approved before putting the

system affected into use.

Validation plan

The plan should first identify:

What is being validated

Where the validation will take place

Why the validation is taking place providing reference to any relevant change.

The validation stages required

Validation time-frames

Overall management of the validation

Performing the validation and recording the outcome

Validation protocol

A validation protocol is an integral element of the validation plan. The protocol describes:

The qualification/validation phase (DQ, IQ, OQ, and PQ)

That tests will be performed

The objectives of the validation in terms of acceptance criteria for each test

Records to be completed.

What needs to be tested, how many tests to do and the acceptance criteria at each validation phase

will be specific to each validation and must be founded on the scientific and technical basis of the

processes and systems involved. It should be possible to establish the specific requirements by

reference to the relevant risk assessments, URS, published standards, regulations & guidelines.



Validation record proformas should be completed as part of the protocol and approved along with

the protocol10

.

THE DIFFERENT TESTS ARE FOLLOWS FOR VALIDATION OF AUTOCLAVE

Vacuum leak test

Status of equipment under validation

Table 1: Status of vacuum leak test

1 Equipment Auto clave cum bung processor

2 Make Machine fabric

3 Capacity 1215 liters

4 Acceptance criteria 123±2 °C

5 Cycle time 121 °C@ 10 min

6 Temperature range 121 °C to 125 °C

7 Pressure range 1.2 bar to 1.3 bar

8 Type of sensor used T type

9 Cycle name Vacuum leak test

Result analysis

Table 2: Probes result of vacuum leak test

Temperature [°C]

Channel Min Max Average Std dev

CH1 121.6 122.7 122.1 0.30

CH2 121.7 123.4 122.4 0.39

CH3 121.6 123.1 122.3 0.39

CH4 122.0 123.4 122.7 0.37

CH5 122.0 123.7 122.8 0.43

CH6 121.9 123.3 122.5 0.37

CH7 121.5 123.3 122.3 0.42

CH8 121.5 123.4 122.3 0.43

CH9 122.3 124.2 123.1 0.48

CH10 121.7 123.2 122.4 0.37

CH11 121.1 122.9 121.8 0.51

CH12 121.4 123.3 122.0 0.46

CH13 121.7 123.3 122.5 0.42

CH14 121.6 123.2 122.4 0.38

CH15 121.1 122.7 121.9 0.45

CH16 121.8 123.2 122.5 0.37

Pressure [bar]

CH17 1.198 1.297 1.262 0.010



Table 3: Result of vacuum leak test

1 Test start time 10:48:06

2 Sterilization start time 10:52:51

3 Sterilization end time 11:02:51

4 Sterilization hold time 10 min

5 Standard sterilization cycle 123±2 °C

6 Time at which first probe reached desired temperature 10:52:21

7 Time at which last probe reached desired temperature 10:52:51

8 Maximum temperature recorded 124.2 °C

9 Minimum temperature recorded 121.1 °C

10 Maximum pressure recorded 1.297 bar

11 Minimum pressure recorded 1.198 bar

12 Test end time 11:04:17

Bowie Dick test


Table 4: Status of Bowie Dick test

1 Equipment Autoclave cum bung processor




5 Cycle time 121 °C @ 15 min

6 Temperature range 121 °C to 125°C



9 Cycle name Bowie dick test

Result analysis



Table 5: Probes result of Bowie Dick test

Temperature [°C]

Probes Min Max Average Std Dev

P1 121.3 122.6 122.0 0.33

P2 121.2 123.7 122.4 0.49

P3 121.1 122.5 121.8 0.35

P4 121.3 123.4 122.1 0.50

P5 121.5 123.1 122.3 0.38

P6 121.4 123.9 122.4 0.52

P7 121.3 122.5 121.5 0.33

P8 121.1 122.6 121.7 0.41

P9 121.4 123.4 122.2 0.50

P10 121.2 122.9 121.5 0.41

P11 121.1 122.6 121.5 0.42

P12 121.3 123.6 122.2 0.55

P13 121.2 123.2 121.9 0.51

P14 121.1 122.4 121.6 0.39

P15 121.3 122.3 121.5 0.32

P16 121.1 122.5 121.7 0.35

Pressure [bar]

P17 1.218 1.360 1.284 0.010

Table 6: Result of Bowie Dick test





5 Standard sterilization cycle 123±2 °C










Graph 1 : Bowie Dick test graph

Figure 2: Results of Bowie Dick test sheets

Heat distribution study

Status of equipment under validation:

Table 7: Status of heat distribution test

1 Equipment Autoclave cum bung processor




5 Cycle time 121 °C @ 30 min

6 Temperature range 121 °C to 125 °C



9 Cycle name Without load



Result analysis

Table 8: Probes result of heat distribution test

Temperature[°C]


CH1 121.1 123.1 121.7 0.35

CH2 121.8 123.6 122.2 0.35

CH3 122.1 123.8 122.7 0.42

CH4 122.1 123.7 122.6 0.37

CH5 121.8 123.6 122.3 0.38

CH6 122.1 123.7 122.5 0.38

CH7 121.8 123.7 122.5 0.41

CH8 121.9 123.7 122.4 0.39

CH9 121.6 123.6 122.1 0.34

CH10 121.7 123.7 122.3 0.42

CH11 122.0 123.8 122.6 0.42

CH12 121.4 123.4 122.0 0.35

CH13 121.7 123.8 122.3 0.50

CH14 121.2 123.2 121.9 0.33

CH15 121.7 123.8 122.3 0.46

CH16 121.5 123.8 122.1 0.41

Pressure [bar]

CH17 1.201 1.292 1.219 0.018

Table 9: Result of heat distribution test





5 Standard sterilization cycle 123 °c±2 °C





10 Maximum pressure recorded 1.292bar

11 Minimum pressure recorded 1.201bar




Result analysis of biological indicator

Table 10 : Biological indicator result of heat distribution study

Sterilization method By steam at a temperature of 121 °C and

pressure of 1.2 bar

Standard sterilization cycle 30 minutes

Biological indicators used Self contained ampoules of Geobacillus

stearothermophilus population 106

cfu/strip

Incubation temperature 55±2 °C

Table 11: Observation of biological indicator

S. No. Position of biological

indicator

Observation

24 hours 48 hours

1 P1 - -

2 P2 - -

3 P3 - -

4 P4 - -

5 P5 - -

6 P6 - -

7 P7 - -

8 P8 - -

9 P9 - -

10 P10 - -

11 P11 - -

12 P12 - -

13 P13 - -

14 P14 - -

15 P15 - -

16 P16 - -

17 Positive control + +

- = No growth + = Shows growth

Figure 3: Results of chemical indicators for heat distribution test



Graph 2: Heat distribution test graph

Heat penetration study


Table 12: Status of heat penetration study (maximum load)

1 Equipment Auto clave cum bung processor



4 Acceptance criteria 123 ± 2°C

5 Cycle time 121°C @ 30 min

6 Temperature range 121°C to 125°C



9 Cycle name Rubber stopper maximum load (60000)



4.2 Result analysis

Table 13: Probes result of heat penetration study (maximum load)

Temperature[°C]


CH1 121.2 121.8 121.4 0.17

CH2 121.3 122.1 121.6 0.19

CH3 121.7 122.8 122.1 0.22

CH4 121.3 122.0 121.6 0.17

CH5 122.1 122.9 122.4 0.20

CH6 121.4 122.0 121.6 0.17

CH7 121.5 122.2 121.8 0.17

CH8 121.5 122.1 121.8 0.15

CH9 122.0 122.7 122.3 0.15

CH10 121.5 122.1 121.7 0.17

CH11 121.4 122.1 121.7 0.17

CH12 121.4 122.1 121.7 0.17

CH13 121.5 122.1 121.8 0.16

CH14 121.5 122.1 121.7 0.17

CH15 121.4 122.0 121.6 0.16

CH16 121.1 122.3 121.6 0.34

Pressure [bar]

CH17 1.202 1.247 1.219 0.011

Table 14: Result of heat penetration study (maximum load)





5 Standard sterilization cycle 123 °C ± 2 °C










Figure 4: Rubber bungs for heat penetration test (maximum load)

Result analysis of biological indicator

Table 15: Biological indicator result of heat penetration study (maximum)

Sterilization method By steam at a temperature of 121 °C and pressure

of 1.2 bar

Standard sterilization cycle 30 minutes

Biological indicators used Self contained ampoules of Geobacillus

stearothermophilus population 106

cfu/strip

Incubation temperature 55±2 °C

Table 16: Observation of biological indicator

S. No. Position of biological

indicator

Observation

24 hours 48 hours

1 P1 - -

2 P2 - -

3 P3 - -

4 P4 - -

5 P5 - -

6 P6 - -

7 P7 - -

8 P8 - -

9 P9 - -

10 P10 - -

11 P11 - -

12 P12 - -

13 P13 - -

14 P14 - -

15 P15 - -

16 P16 - -

17 Positive control + +

- = No growth + = Shows growth



Figure 5: Results of chemical indicators for heat penetration study (maximum load)

Graph 3: Heat penetration test (maximum load) graph

CONCLUSION

After conducting the Vacuum leak test, Bowie-dick test, Heat distribution test, Heat penetration

test, the results are satisfied and results are captured in the above respective tables and hence it

proves that equipment meets its acceptance criteria and equipment passes the above tests.

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